High Red–Blue Light Ratio Promotes Accelerated In Vitro Flowering and Seed-Set Development in Amaranthus hypochondriacus Under a Long-Day Photoperiod

Grain amaranths are recalcitrant to conventional in vitro plant regeneration by organogenesis de novo or through somatic embryogenesis. Consequently, floral organogenesis by these methods, representing the culminating developmental point in angiosperms, is rarely achieved. In the present study, the manipulation of in vitro flowering was explored as part of a strategy designed to overcome grain amaranth's regeneration recalcitrance. It led to an efficient and reproducible in vitro protocol in which half-longitudinally dissected zygotic embryos generated fully developed <i>Amaranthus hypochondriacus</i> (<i>Ah</i>) plants. The use of high-irradiance illumination with LED lamps with a 3:1 red-blue irradiance ratio was a critical factor, leading to a 70% rate of early flowering events under flowering-inhibiting long-day photoperiod conditions. Contrariwise, no flowering was induced under LED white lights. All in vitro flowering <i>Ah</i> plants yielded viable seeds. To understand the basic molecular mechanisms of the phenomenon observed, gene expression patterns and principal component analysis of key flowering-related genes were analyzed after cultivation in vitro for 4, 8, and 12 weeks under both lighting regimes. These coded for photoreceptors, photomorphogenetic regulators, embryogenic modulators, and flowering activators/repressors. The results highlighted the upregulation of key flowering-regulatory genes, including <i>CONSTANS</i>, <i>FLOWERING LOCUS T</i>, and <i>LEAFY</i>, together with the downregulation of the floral repressor <i>TERMINAL FLOWER1</i>. Ribosome biogenesis- and seed-development-related genes were also differentially expressed, supporting a key role in this process for protein synthesis and embryogenesis. A model is proposed to explain how this light-regulated molecular framework enables in vitro flowering and seed production in <i>Ah</i> plants kept under long-day photoperiods.